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The RNA-binding motif 45 (RBM45) protein accumulates in inclusion bodies in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with TDP-43 inclusions (FTLD-TDP) patients.

Collins M, Riascos D, Kovalik T, An J, Krupa K, Krupa K, Hood BL, Conrads TP, Renton AE, Traynor BJ, Bowser R - Acta Neuropathol. (2012)

Bottom Line: These RBM45 inclusions were observed in spinal cord motor neurons, glia and neurons of the dentate gyrus.By confocal microscopy, RBM45 co-localizes with ubiquitin and TDP-43 in inclusion bodies.In neurons containing RBM45 cytoplasmic inclusions we often detected the protein in a punctate pattern within the nucleus that lacked either TDP-43 or ubiquitin.

View Article: PubMed Central - PubMed

Affiliation: Departments of Neurobiology and Pathology, University of Pittsburgh, PA, USA.

ABSTRACT
RNA-binding protein pathology now represents one of the best characterized pathologic features of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration patients with TDP-43 or FUS pathology (FTLD-TDP and FTLD-FUS). Using liquid chromatography tandem mass spectrometry, we identified altered levels of the RNA-binding motif 45 (RBM45) protein in the cerebrospinal fluid (CSF) of ALS patients. This protein contains sequence similarities to TAR DNA-binding protein 43 (TDP-43) and fused-in-sarcoma (FUS) that are contained in cytoplasmic inclusions of ALS and FTLD-TDP or FTLD-FUS patients. To further characterize RBM45, we first verified the presence of RBM45 in CSF and spinal cord tissue extracts of ALS patients by immunoblot. We next used immunohistochemistry to examine the subcellular distribution of RBM45 and observed in a punctate staining pattern within nuclei of neurons and glia in the brain and spinal cord. We also detected RBM45 cytoplasmic inclusions in 91 % of ALS, 100 % of FTLD-TDP and 75 % of Alzheimer's disease (AD) cases. The most extensive RBM45 pathology was observed in patients that harbor the C9ORF72 hexanucleotide repeat expansion. These RBM45 inclusions were observed in spinal cord motor neurons, glia and neurons of the dentate gyrus. By confocal microscopy, RBM45 co-localizes with ubiquitin and TDP-43 in inclusion bodies. In neurons containing RBM45 cytoplasmic inclusions we often detected the protein in a punctate pattern within the nucleus that lacked either TDP-43 or ubiquitin. We identified RBM45 using a proteomic screen of CSF from ALS and control subjects for candidate biomarkers, and link this RNA-binding protein to inclusion pathology in ALS, FTLD-TDP and AD.

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TDP-43 pathology can occur independent of RBM45 pathology in ALS and FTLD cases. RBM45 is labeled in green, TDP-43 denoted in red and DAPI (blue) visualizes nuclei in the merged images. a Motor neuron with nuclear RBM45 and a TDP-43 positive inclusion that labels poorly for RBM45. Several such inclusions were found throughout the lumbar spinal cord of sALS and fALS cases. b Motor neurons from C9ORF72 ALS cases exhibited nuclear depletion of RBM45. RBM45 was not contained in the TDP-43 inclusions in this motor neuron. c Several TDP-43 positive, RBM45 negative inclusions are indicated by arrowheads in the dentate gyrus of an FTLD case. While no RBM45 positive inclusions were seen, the speckled nuclear staining pattern was observed in several adjacent cells. Scale bar 20 μm. Panels represent the following case numbers in Table 1: a 7, b 2, c 27
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Fig6: TDP-43 pathology can occur independent of RBM45 pathology in ALS and FTLD cases. RBM45 is labeled in green, TDP-43 denoted in red and DAPI (blue) visualizes nuclei in the merged images. a Motor neuron with nuclear RBM45 and a TDP-43 positive inclusion that labels poorly for RBM45. Several such inclusions were found throughout the lumbar spinal cord of sALS and fALS cases. b Motor neurons from C9ORF72 ALS cases exhibited nuclear depletion of RBM45. RBM45 was not contained in the TDP-43 inclusions in this motor neuron. c Several TDP-43 positive, RBM45 negative inclusions are indicated by arrowheads in the dentate gyrus of an FTLD case. While no RBM45 positive inclusions were seen, the speckled nuclear staining pattern was observed in several adjacent cells. Scale bar 20 μm. Panels represent the following case numbers in Table 1: a 7, b 2, c 27

Mentions: In the hippocampus of FTLD-TDP and AD patients, RBM45 co-localized with TDP-43 in cytosolic and rare intranuclear inclusions of dentate granule cells (arrows in Fig. 5d, e). We also observed co-localization of RBM45 to TDP-43 inclusions in the dentate of AD patients (Fig. 5f). A speckled RBM45 nuclear staining pattern was evident in many dentate granule cells that is distinct from the more diffuse TDP-43 nuclear immunostaining. Using multiple images from 10 ALS, 4 FTLD-TDP and 5 AD cases, we determined that RBM45 was present in 64 % of the TDP-43 inclusions in the spinal cord of ALS cases and 70 % of the TDP-43 inclusions in the hippocampus of FTLD-TDP and AD cases. Examples of TDP-43 inclusions that lack RBM45 are shown in Fig. 6. We observed ALS spinal cord motor neurons with nuclear RBM45 and weak or absent co-localization to TDP-43 cytoplasmic inclusions (Fig. 6a). In subjects with the C9ORF72 repeat expansion, we also observed spinal cord motor neurons devoid of nuclear RBM45 and lacking RBM45 positive cytoplasmic inclusions (Fig. 6b). Finally, the hippocampus of ALS, FTLD-TDP and AD subjects contained TDP-43 inclusions without RBM45 (arrowheads in Fig. 6c). We did not detect RBM45 inclusions that completely lack TDP-43 immunoreactivity.Fig. 6


The RNA-binding motif 45 (RBM45) protein accumulates in inclusion bodies in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with TDP-43 inclusions (FTLD-TDP) patients.

Collins M, Riascos D, Kovalik T, An J, Krupa K, Krupa K, Hood BL, Conrads TP, Renton AE, Traynor BJ, Bowser R - Acta Neuropathol. (2012)

TDP-43 pathology can occur independent of RBM45 pathology in ALS and FTLD cases. RBM45 is labeled in green, TDP-43 denoted in red and DAPI (blue) visualizes nuclei in the merged images. a Motor neuron with nuclear RBM45 and a TDP-43 positive inclusion that labels poorly for RBM45. Several such inclusions were found throughout the lumbar spinal cord of sALS and fALS cases. b Motor neurons from C9ORF72 ALS cases exhibited nuclear depletion of RBM45. RBM45 was not contained in the TDP-43 inclusions in this motor neuron. c Several TDP-43 positive, RBM45 negative inclusions are indicated by arrowheads in the dentate gyrus of an FTLD case. While no RBM45 positive inclusions were seen, the speckled nuclear staining pattern was observed in several adjacent cells. Scale bar 20 μm. Panels represent the following case numbers in Table 1: a 7, b 2, c 27
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Fig6: TDP-43 pathology can occur independent of RBM45 pathology in ALS and FTLD cases. RBM45 is labeled in green, TDP-43 denoted in red and DAPI (blue) visualizes nuclei in the merged images. a Motor neuron with nuclear RBM45 and a TDP-43 positive inclusion that labels poorly for RBM45. Several such inclusions were found throughout the lumbar spinal cord of sALS and fALS cases. b Motor neurons from C9ORF72 ALS cases exhibited nuclear depletion of RBM45. RBM45 was not contained in the TDP-43 inclusions in this motor neuron. c Several TDP-43 positive, RBM45 negative inclusions are indicated by arrowheads in the dentate gyrus of an FTLD case. While no RBM45 positive inclusions were seen, the speckled nuclear staining pattern was observed in several adjacent cells. Scale bar 20 μm. Panels represent the following case numbers in Table 1: a 7, b 2, c 27
Mentions: In the hippocampus of FTLD-TDP and AD patients, RBM45 co-localized with TDP-43 in cytosolic and rare intranuclear inclusions of dentate granule cells (arrows in Fig. 5d, e). We also observed co-localization of RBM45 to TDP-43 inclusions in the dentate of AD patients (Fig. 5f). A speckled RBM45 nuclear staining pattern was evident in many dentate granule cells that is distinct from the more diffuse TDP-43 nuclear immunostaining. Using multiple images from 10 ALS, 4 FTLD-TDP and 5 AD cases, we determined that RBM45 was present in 64 % of the TDP-43 inclusions in the spinal cord of ALS cases and 70 % of the TDP-43 inclusions in the hippocampus of FTLD-TDP and AD cases. Examples of TDP-43 inclusions that lack RBM45 are shown in Fig. 6. We observed ALS spinal cord motor neurons with nuclear RBM45 and weak or absent co-localization to TDP-43 cytoplasmic inclusions (Fig. 6a). In subjects with the C9ORF72 repeat expansion, we also observed spinal cord motor neurons devoid of nuclear RBM45 and lacking RBM45 positive cytoplasmic inclusions (Fig. 6b). Finally, the hippocampus of ALS, FTLD-TDP and AD subjects contained TDP-43 inclusions without RBM45 (arrowheads in Fig. 6c). We did not detect RBM45 inclusions that completely lack TDP-43 immunoreactivity.Fig. 6

Bottom Line: These RBM45 inclusions were observed in spinal cord motor neurons, glia and neurons of the dentate gyrus.By confocal microscopy, RBM45 co-localizes with ubiquitin and TDP-43 in inclusion bodies.In neurons containing RBM45 cytoplasmic inclusions we often detected the protein in a punctate pattern within the nucleus that lacked either TDP-43 or ubiquitin.

View Article: PubMed Central - PubMed

Affiliation: Departments of Neurobiology and Pathology, University of Pittsburgh, PA, USA.

ABSTRACT
RNA-binding protein pathology now represents one of the best characterized pathologic features of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration patients with TDP-43 or FUS pathology (FTLD-TDP and FTLD-FUS). Using liquid chromatography tandem mass spectrometry, we identified altered levels of the RNA-binding motif 45 (RBM45) protein in the cerebrospinal fluid (CSF) of ALS patients. This protein contains sequence similarities to TAR DNA-binding protein 43 (TDP-43) and fused-in-sarcoma (FUS) that are contained in cytoplasmic inclusions of ALS and FTLD-TDP or FTLD-FUS patients. To further characterize RBM45, we first verified the presence of RBM45 in CSF and spinal cord tissue extracts of ALS patients by immunoblot. We next used immunohistochemistry to examine the subcellular distribution of RBM45 and observed in a punctate staining pattern within nuclei of neurons and glia in the brain and spinal cord. We also detected RBM45 cytoplasmic inclusions in 91 % of ALS, 100 % of FTLD-TDP and 75 % of Alzheimer's disease (AD) cases. The most extensive RBM45 pathology was observed in patients that harbor the C9ORF72 hexanucleotide repeat expansion. These RBM45 inclusions were observed in spinal cord motor neurons, glia and neurons of the dentate gyrus. By confocal microscopy, RBM45 co-localizes with ubiquitin and TDP-43 in inclusion bodies. In neurons containing RBM45 cytoplasmic inclusions we often detected the protein in a punctate pattern within the nucleus that lacked either TDP-43 or ubiquitin. We identified RBM45 using a proteomic screen of CSF from ALS and control subjects for candidate biomarkers, and link this RNA-binding protein to inclusion pathology in ALS, FTLD-TDP and AD.

Show MeSH
Related in: MedlinePlus